45 A HIGHLY SENSITIVE AND SPECIFIC SYSTEM FOR LARGE-SCALE GENE EXPRESSION PROFILING. (1st March 2007)

• Record Type: Journal Article
• Authors: Hu, G.; Yang, Q.; Li, H.
• Is Part Of: Journal of investigative medicine. Volume 55:Number 2(2007)
• Page Start: S356
• Page End: S356
• Publication Date: 2007-03-01
• Subjects: Clinical medicine -- Periodicals; Medicine -- Research -- Periodicals; Medicine; Research -- United States; Clinical medicine; Medicine -- Research; Periodicals; 616.075
• DOI: 10.1136/jim-55-02-45
• Languages: English
• ISSNs: 1081-5589

Abstract

Abstract : Gene expression profiling of very few or even single cells is of particular interest in many applications, including molecular diagnosis. However, detection of a large number of mRNA sequences from a small number of cells is limited by the sensitivity of available methods. High-throughput multiplex reverse transcription followed by PCR amplification (RT-PCR) has much to offer these studies owing to its inherent sensitivity, efficiency, and cost-effectiveness. A multiplex RT-PCR-based high-throughput gene profiling system is described in this communication. With this system, a breast cancer-specific microarray is developed. More than 1, 000 different mRNA species can be amplified in a single tube to a detectable amount. By using specially designed PCR primers, the long-standing low-specificity problem associated with high-throughput gene expression profiling has been solved. Amplified sequences are then resolved by microarray with probes that only hybridize to sequences amplified from mRNA. The method is so sensitive that mRNA in single cells can be reliably detected. Differentially expressed genes identified with the high-throughput approach in the breast cancer cell line MCF-7 and its drug-resistant variant MCF-7/AdrR could be validated by a different method. The approach may greatly facilitate the analysis of combinatorial expression of known genes in many important applications with a limited amount of RNA, such as molecular diagnosis of cancer patient with samples of fine-needle aspiration prior to surgery.